Quantitative aspects of the effects of insulin, epidermal growth factor and dexamethasone on DNA synthesis in cultured adult rat hepatocytes.

Epidermal growth factor (EGF) and insulin in combination have previously been shown to initiate S-phase in primary cultures of adult rat hepatocytes. We here describe the detailed time course and dose-dependency of the effects of EGF and insulin on DNA synthesis in cultured hepatocytes. The DNA synthesis was assessed either biochemically or autoradiographically with a fairly good correlation between the two methods. DNA synthesis started 24-30 h after plating of the cells and peaked at approximately 70 h. Up to 70% of the cells entered DNA synthesis during this period. EGF and insulin acted synergistically on the DNA synthesis. Dexamethasone raised the DNA synthesis slightly, maximal effect occurred at concentrations above 2.5 nM and this agent was routinely used in the experiments with EGF and insulin. In the presence of 0.4 microM insulin from the time of plating, EGF dose-dependently increased the DNA synthesis with maximal effect at 5-15 nM. When added in combination with 1.7 nM EGF, insulin enhanced the DNA synthesis over the concentration range from 0.1 to 3 nM. These studies show that primary cultures of hepatocytes are useful in assessing the quantitative aspects of the interactions between the growth stimulating effects of hormones.